Cercarial productivity of redial generations in single-miracidium infections of Lymnaea truncatula with Paramphistomum daubneyi or Fasciola hepatica

Single-miracidium infections of Lymnaea truncatula with Paramphistomum daubneyi or with Fasciola hepatica were carried out under laboratory conditions to count free rediae, their germinal embryos, and to determine the cercarial productivity of each redial generation. In snails infected by P. daubneyi, the cercariae were produced by the first (8.7 cercariae per redia) and second (8.9 per redia) generations. At day 63 post-exposure, they corresponded, respectively, to 53.9% and 46.1% of cercariae produced by all rediae. In snails infected by F. hepatica, the majority of cercariae were produced by the R2a group (18.2 cercariae per redia) and corresponded to 66.0% of cercariae produced all rediae. The cercariae produced by the other redial groups were more limited in number: 17.5 per redia in the R1b group (28.7%) and 2.0 per redia in the R2b/R3a group (5.3%). Cercarial productivity of P. daubneyi until day 63 post-exposure was more limited in number than that of F. hepatica: a total of 145 cercariae per snail versus 427 per snail.     

Fasciola hepatica: the growth and larval productivity of redial generations in Galba truncatula subjected to miracidia differing in their mammalian origin

Experimental infections of Galba truncatula with 4 isolates of Fasciola hepatica miracidia differing by their mammalian origin (cattle, nutrias, rabbits, or sheep) were carried out to determine if parasite origin had an effect on the number of free rediae, their growth, and their larval productivity in each redia category. The mammalian origin of miracidia had a significant influence on the numbers of free rediae (they were higher in cattle-group snails) and the lengths of rediae (they were lower in rabbit groups). The redia category had also a significant effect on body and pharyngeal measurements. In all groups, the majority of cercariae (55.8-63.2%) were produced by the daughter rediae (R2a rediae) originating from the first mother redia. Compared with the other groups, the mean number of cercariae at day 49 postexposure was twice as high in cattle groups. In contrast, the mean number of daughter rediae produced by each second-appearing mother redia or each R2a redia was higher in the nutria, rabbit, and sheep groups. The mammalian origin of F. hepatica miracidia had an effect on the number of live rediae, their length, and their redial and cercarial productivity.     

Insights into the development of Notocotylus attenuatus (Digenea: Notocotylidae) in Lymnaea stagnalis: from mother sporocyst to cercariae

Notocotylus attenuatus (Digenea: Notocotylidae) is a monostome fluke parasitizing the intestinal caeca of waterfowl that uses an injection apparatus to infect its intermediate snail host. Morphology of the invading larva (a sporocyst), and the intramolluscan larval development of this fluke have not been characterized extensively. In this study, experimental infections of Lymnaea stagnalis using N. attenuatus eggs resulted in the development of sporocysts containing one germ ball or mother redia between 12 and 21 days post exposure (p.e.) within the hepatopancreas. Independent mother rediae and developing daughter rediae were present between day 25 and day 42 p.e. Cercariae, within the body of rediae, were detected 42 days p.e. The development of daughter rediae and cercariae started posteriorly in the body of parent redia and these larvae migrated anteriorly during development towards the birth pore. A cercaria was also observed emerging from the birth pore and released cercariae maturated further within the snail hepatopancreas prior to leaving the snail. The intramolluscan development was completed 45 days p.e. when the first fully formed cercariae were shed into the outer environment. These data detail the fascinating post-embryonic development of N. attenuatus and highlight the intricate nature of larval transitions within its snail host.     

Histological study of the development of Fasciola hepatics in Lymnaea truncatula, L. glabra and L. palustris infested at hatching]

The development of Fasciola hepatica is histologically analysed in Limnaea truncatula, L. glabra and L. palustris infested at hatching with a single miracidium. The development of the redial generations of Fasciola is not affected by the species of snail host but depends on growth of the snail. In snails of size within 2 mm (considered at the day 49 of infestation to 20 degrees C), the rediae show a delay of maturity compared with rediae observed in snails of higher sizes. The distribution of redia 1 of generation 1 should have an influence on growth of the snail host.     

Redial generations of Fasciola gigantica in the pulmonate snail Lymnaea truncatula.

Lymnaea truncatula, 4 mm in height, were subjected to infection by a single miracidium of Fasciola gigantica, then raised at 23 degrees C until day 60 of the experiment. Histological study of these snails demonstrated a mean redial burden of 34 parasites at day 60, of which one third were degenerating forms. The mean number of living independent rediae did not exceed 5 for the first and second generations. Conversely, in subsequent generations there were as many as 18 rediae per snail at day 60. The first living redia of the first generation in particular gave rise to daughter rediae. Mature rediae appeared at day 35 and especially concerned the first and second generations at day 60. The authors conclude that development of the first and second redial generations occurs during the same period, and that the forms of the first cohort of the second generation are produced from the first redia of the first generation which originated from the sporocyst.     

Observations on the host-parasite relations between Echinostoma revolutum and lymnaeid snails.

Echinostoma revolutum from Taiwan was studied in lymneid snails at 29 +/- 0.5 degrees C. Given 3-5 miracidia, 95% of Lymnaea ollula and 40% of Lymnaea swinhoei became positive; the prepatent periods were 18 and 25 days, respectively. The following are based on the observations in Lymnaea ollula: The time required for miracidial penetration was about one hour. The sporocysts developed only in the ventricle of the snail but mother rediae developed in the heart and other organs. Mature daughter rediae were not found in the heart cavity. The sporocysts reached the ventricle within 3 days. Mother rediae were released after 6 days and daughter rediae after 8 days. Given 5 miracidia, 1-3 sporocysts reached the heart and 2-20 mother rediae were found per snail. The number of mature daughter rediae was usually 100-200 although more than a thousand may develop in a snail. The sporocysts and mother rediae attained maximal size 9 days postinfection and started degeneration 13 days postinfection. Daughter rediae were largest at the beginning of cercarial emergence and decreased in size thereafter. Simultaneous production of daughter rediae and cercariae by the mother redia was seen only once in this snail mature cercariae were obtained in 10 days postinfection. The cercariae emerged from a small area of mantle collar near the posterior corner of shell aperture. They were negatively phototactic and positively geotactic. An estimation showed that each snail shed about 350 cercariae a day. The cercariae reached the pericardial cavity of snail in one hour via the renal orifice and metacercariae were seen 4-5 hours after exposure. The infectivity of cercariae at various times after shedding, as expressed by cyst recovery rates, were: 51.6%, O-hr old; 76.1%, 2-hr; 68% 4-hr; 32%, 6-hr; 3%, 8 and 10-hr. Cyst recovery rates were not different within the dosage of 50-500 cercariae per snail. Most metacercariae recovered 1-2 days after cercarial exposure were viable; only 5 among 6,533 cysts were dead.     

Fasciola hepatica: cercarial productivity of redial generations in long-surviving Galba truncatula

Bimiracidial infections of Galba truncatula with Fasciola hepatica were carried out to determine the effect of food quality on the frequency of 1- and 2-sporocyst infections, to analyse its impact on the developmental patterns (normal, or abnormal) of redial generations, and to verify its consequences on cercarial production. These investigations were performed in snails reared at 20 degrees C and provided with cos lettuce and commercial fish food (Tetraphyll) as a food source until their death. Double-sporocyst infections with normal development of redial generations were recorded in 43.9% of infected snails (out of 296). Single-sporocyst infections were noted in the other snails, with normal development of generations in 53.7% and abnormal development (the first mother redia early degenerated) in 2.4%. Four successive redial generations were found in long-surviving snails (more than 90 days). In both 1- and 2-sporocyst infections, showing normal development of generations, the daughter rediae, which exited from the first mother redia (R2a rediae), constituted the greater group of free rediae and produced the highest percentages of cercariae (46.2-48.2%). However, the development of these rediae inside the snail body was slower in 2-sporocyst infections than in 1-sporocyst infections. The numbers of rediae noted in subsequent generations (R2b/R3a and R3b/R4a rediae) were similar, whatever the number of full-grown sporocysts. The number of shed cercariae recorded in the 1- and 2-sporocyst infections did not significantly differ. When long-surviving snails died, 19.8-20.7% of cercariae produced by free rediae (essentially by R2b/R3a and R3b/R4a rediae) were still present in their bodies. The increased frequency of 2-sporocyst infections demonstrated that food quality had a significant effect on the redial burden of F. hepatica developing inside G. truncatula.     

In vitro culture of the avian echinostome Himasthla elongata: from redia to marita

Axenic primary cultures of Himasthla elongata rediae harvested from hepatopancreas of naturally infected marine prosobranch snail Littorina littorea were maintained in Leibovitz's L-15 medium (osmolarity of approximately 780 mOsm, pH 7.8, temperature 14 degrees C under normal atmospheric conditions). Cultured rediae were active, motile and demonstrated high synthetic activity in metabolic labelling experiment. Long-term cultivation experiment showed 50% survival level of the rediae for up to 70 days and significant differences between mortality in redia groups derived from different host individuals. Half of the rediae in the most robust group survived for up to 163 days, when the experiment was terminated. Development and emergence of in vivo preformed cercariae and daughter rediae was observed. Cercariae in the culture also encysted, transformed into metacercariae and some of them in one to two weeks after the transformation spontaneously excysted into juvenile maritae. The employed culture system is characterized by a very low level of proteolytic activity. This system is suggested as a method permitting to obtain rediae secretory-excretory products free of host-derived contaminants.     

The life-cycle of Petasiger islandicus Kostadinova & Skirnisson, 2007 (Digenea: Echinostomatidae) elucidated with the aid of molecular data

The small planorbid snail Gyraulus cf. laevis (Alder) from Lake Myvatn in Iceland was found to emit large-tailed cercariae with 19 collar spines, and three-spined sticklebacks Gasterosteus aculeatus L. were infected with metacercariae of a species of Petasiger Dietz, 1909. Comparative sequence analysis using ND1 mitochondrial DNA sequences revealed that the rediae and cercariae are conspecific with P. islandicus Kostadinova & Skirnisson, 2007, recently described from an isolated population of the horned grebe Podiceps auritus (L.) at the lake. The redia, cercaria and metacercaria are described and compared with related forms.     

Hyperparasitism by Paragordius varius (Nematomorpha: Gordiida) larva of Monostome redia (Trematoda: Digenea)

Gordiid larvae enter and encyst within a large variety of aquatic vertebrates and invertebrates. Cysts of a single species of gordiid have been found in such diverse hosts as insect larvae, snails, crustaceans, fish, and many others. One particularly surprising host report was made nearly 100 yr ago, i.e., cysts within adult trematodes. The origin of these cysts remained unclear, but it was suggested that the gordiid larvae gained entry into the adult flukes via their definitive, vertebrate host. In the present study, the finding is reported of gordiid cysts within monostome redia within a Physa gyrina snail. This is the first time gordiid cysts have been reported within redia, and this provides an alternative explanation for the presence of cysts within adult trematodes.     

Experimental Life History and Biological Characteristics of Fasciola gigantica (Digenea: Fasciolidae)

This study was conducted to investigate the life history, morphology, and maturation of larval stages and adult worms of Fasciola gigantica in experimental mice. Lymnaea auricularia rubiginosa was used as the intermediate host, and Oryza sativa was used for encystment of the metacercariae, while Mus musculus was used as the definitive host for maturation study. Fresh eggs from the gall bladder of water buffaloes fully developed into embryonated ones and hatched out at days 11-12 after incubation at about 29ºC. Free-swimming miracidia rapidly penetrated into the snail host, and gradually developed into the next larval stages; sporocyst, redia, and daughter redia with cercariae. Fully-developed cercariae were separated from the redia and shed from the snails on day 39 post-infection (PI). Free-swimming cercariae were immediately allowed to adhere to rice plants, and capsules were constructed to protect metacercariae on rice plants. Juvenile worms were detected in intestines of mice at days 3 and 6 PI, but they were found in the bile duct from day 9 PI. Juvenile and adult flukes were recovered from 16 mice experimentally infected with metacercariae, with the average recovery rate of 35.8%. Sexually mature adult flukes were recovered from day 42 PI. It could be confirmed that experimentally encysted metacercariae could infect and develop to maturity in the experimental host. The present study reports for the first time the complete life history of F. gigantica by an experimental study in Thailand. The obtained information can be used as a guide for prevention, elimination, and treatment of F. gigantica at environment and in other hosts.     

Cercarial shedding of Echinostoma cinetorchis and experimental infection of the cercariae to several kinds of snails

The development of Echinostoma cinetorchis in several snail species reared in laboratory aquaria was observed. The eggs from adult flukes collected from the intestine of rats were cultivated to miracidia, and exposed to Hippeutis sp. snails. Observations were made for cercarial shedding from the exposed snails. The cercariae shed from the snails were again exposed to several species of fresh water snails in order to observe metacercarial formation in the snails and their infectivity to final hosts. The results obtained in this study were as follows: 1. Twenty miracidia were exposed to each snail of Hippeutis sp. About 58.3% of the above snails (7 out of 12) were dead before shedding the cercariae, and the remainder shed the cercariae for a period of 7 to 9 days before death. 2. Cercarial shedding from the infected snails started from the 25th day after the exposure to miracidia, and the total number of cercariae shed per snail was 684 in average (range; 482-904). 3. The size of rediae developed in the infected Hippeutis sp. snails was 1,242 x 214 microns in average, and the number of rediae per snail was 350 in average (range; 120-510). 4. About 40 to 50 cercariae shed from the Hippeutis sp. snails were each exposed to several species of snails reared in the laboratory. The metacercarial formation was confirmed by dissecting the infected snails, 12 to 16 days after the infection.(ABSTRACT TRUNCATED AT 250 WORDS)     

Germinal elements and their development in Echinostoma caproni and Echinostoma paraensei (Trematoda) miracidia.

Among the large cells located in the posterior of Echinostoma caproni and E. paraensei miracidia are secretory cells, germinal cells (GC), and undifferentiated cells. Secretory cells do not give rise to progeny, whereas GC do. Undifferentiated cells develop into GC that can also divide to produce embryos. Cleavage of GC of E. caproni occurs only after the parasite has entered the snail host and develops into a sporocyst. With E. paraensei, GC are larger than noted for E. caproni, and in 3 of 23 miracidia examined, germinal cell cleavage had occurred in the miracidium such that an embryo containing 20-25 blastomeres was present. Observations on the germinal elements of miracidia help to explain previous results showing that (1) E. paraensei sporocysts release mother rediae a few days earlier than do sporocysts of E. caproni, and that (2) a single mother redia is produced ahead of all others by sporocysts of E. paraensei, but not by sporocysts of E. caproni. The present study adds support to the concept that E. caproni and E. paraensei are distinct species.     

Neutral lipids in cercariae, encysted metacercariae, and rediae of Zygocotyle lunata

High-performance thin-layer chromatography was used to analyze the neutral lipids in the rediae, cercariae, and encysted metacercariae of the paramphistomid trematode Zygocotyle lunata. Visual observations of the chromatograms showed that the most abundant lipid fractions were free sterols and free fatty acids in all larval stages and triacylglycerols in the metacercariae and rediae. The weight of free sterols (x +/- SE) was 120+/-20 ng/cercaria, 56+/-3.8 ng/redia, and 5.9+/-1.5 ng/encysted metacercaria; the weight of triacylglycerols was 13+/-0.88 ng/encysted metacercaria, 6.3+/-0.063 ng/redia, and was not detectable in the cercaria; the weight of free fatty acids was 160+/-17 ng/ cercaria, 76+/-9.1 ng/redia, and 4.2+/-0.46 ng/encysted metacercaria. Oil red O staining of whole larvae showed the presence of neutral lipids in the rediae but not in the cercariae or encysted metacercariae. A dramatic reduction was seen in the quantity of free sterols and free fatty acids in the encysted metacercariae as compared with the cercariae, suggesting that these neutral lipids are used in some way during the transformation from cercaria to metacercaria.     

Redial growth and cercarial productivity of Fasciola hepatica in three species of young lymnaeid snails

Experimental infections of 1-mm high snails using three populations of Lymnaea (L. glabra, L. ovata and L. truncatula) and a cattle strain of Fasciola hepatica miracidia were carried out under laboratory conditions to determine if the snail species had an effect on the number of free rediae, their growth, and cercarial productivity in relation to each redial category (R1a, R1b, R2a, or R2b/R3a). The total number of rediae ranged from 6.4 to 7.5 per snail. The mean body length of rediae varied from 1-1.2 mm (R1a) to 0.3-0.4 mm (R2b/R3a). The width of the intrapharyngeal lumen also varied from 26.0-38.8 microm to 3.0-4.2 microm, respectively. The redial category had a significant effect on both measurements, whereas snail species only had a significant influence on body length. The mean number of cercariae produced by all living rediae at day 49 post-exposure ranged from 63.0 in L. glabra to 87.2 in L. truncatula. In L. ovata and L. truncatula, 55.8% and 58.6% of cercariae, respectively, were produced by R2a rediae, whereas 53.9% of cercariae in L. glabra were formed by the R1b rediae. When young snails were infected with F. hepatica, the species of snail had an effect on the number of living rediae, their length and their cercarial productivity.     

Ribeiroia marini: Surface ultrastructure of redia,cercaria, and adult

Rediae, cercariae, and adults of Ribeiroia marini were examined using a scanning electron microscope to determine the types of tegumental sensory structures and their locations. Sensory structures were observed among numerous tegumental folds in the area immediately surrounding the mouth of the rediae. These sensory structures are similar in appearance, location and fine structure to sensory structures described from the anterior tips of rediae known to be predacious on the sporocysts of Schistosoma mansoni. These uniciliated structures may function as chemoreceptors to aid the redia in migration through snail tissue. Five types of sensory structures bearing one, two, or multiple cilia were distinguishable on the cercariae. These structures were located on and around the oral sucker, dorsal and ventral body surfaces and on the tail. They may be used by the cercariae to locate the intermediate host fish and to find suitable sites within the lateral line scales for encystment. The ventral surface of the adult fluke is covered with spines and shows an absence of sensory structures on the general body surface. Sensory structures were seen in the area surrounding the oral and ventral suckers. The extended cirrus organ has a folded tegument, but lacks spines or sensory structures.     

An experimental study of the effects of Nosema eurytremae (Microsporida: Nosematidae) on the liver fluke Fasciola hepatica

A microsporidian parasite, Nosema eurytremae, was fed to Lymnaea trunculata infected with Fasciola hepatica. Microsporidian infection of the snail was always light, though spores were present in all tissues but the rediae became heavily infected in the parenchyma. The proportion of infected rediae increased with time and was especially high in ageing infections. Although rediae were only lightly infected when examined before cercarial release began, the infections became progressively heavier, so that towards the end of the life of the snails a high proportion of rediae were totally destroyed and contained no cercariae. Cercarial output was significantly depressed over a long period of active shedding by snails which had been hyperinfected with 1 x 106 spores and some of the liberated cercariae were themselves infected.     

Aroapyrgus alleei Morrison, 1946 (Gastropoda: Hydrobiidae) first intermediate host of paragonimus mexicanus in Colima, Mexico

The lung fluke Paragonimus mexicanus infects opossums in the state of Colima, on the Pacific coast of Mexico. Some of the hydrobiid snails occurring in the same area, and identified as Aroapyrgus alleei Morrison, 1946, were found naturally infected with rediae and cercariae of P. mexicanus. The morphology of A. alleei is here described and compared to that of other species of Aroapyrgus, which are actual or potential hosts of lung flukes in other enzootic or endemic areas in Central and South America.     

Oxidative enzymes in the development of Fasciola hepatica L. IV. The activity of oxidases and dehydrogenases in redia.

The object of the study was to investigate the occurrence and localization of oxidative enzymes in the redia -- the third larval stage of Fasciola hepatica L. The author detected cytochrome oxidase, peroxidase, NADH and NADPH tetrazolium reductases (diaphorases), as well as succinate, isocitrate, malate, lactate, alpha-glycerophosphate, glyceraldehyde phosphate, glucose-6-phosphate, 6-phosphogluconate, beta-hydroxybutyrate, L-glutamate, and alcohol dehydrogenases. The presence and localization of the enzymes in various periods of development of the redia were detected with histochemical methods. Out of the studied oxidases and dehydrogenases only cytochrome oxidase was found to be absent from the stages of young rediae. It was ascertained that the redia uses all three paths of release of energy i.e. the glycolytic, Krebs, and pentose cycles, glycolysis being presumably the principal mode of energy production.     

Larval development of Fasciola hepatica in experimental infections: variations with populations of Lymnaea truncatula

A retrospective study was undertaken on 70 French populations of Lymnaea truncatula experimentally infected with Fasciola hepatica to determine whether or not susceptibility of snails to infection influenced redial and cercarial production. Results were compared with those obtained from two control populations, known for prevalences higher than 60% when experimentally infected with F. hepatica. In the 70 other populations examined, the prevalences ranged from 2 to 75%. In 55 of these populations, where the prevalence was more than 20%, a high proportion (50.1-56.8%) of snails died after cercarial shedding, whereas in the other groups (non-shedding snails with the most differentiated larvae being free cercariae, rediae containing cercariae, immature rediae, or sporocysts, respectively), snail death was significantly less. In 11 populations, where the prevalence values were 5-19%, only 14% of snails died after cercarial shedding, whereas snails with free cercariae, rediae with cercariae, or immature rediae showed significant increases in snail mortality. In the remaining four snail populations, with prevalences of less than 5%, the most differentiated larval forms were only immature rediae and/or sporocysts. Overall, the number of rediae containing cercariae significantly decreased with decreasing prevalence values. The low prevalence of experimental infection in several populations of snails might be explained by the occurrence of natural infections with miracidia originating from a mammalian host other than cattle, and/or by genetic variability in the susceptibility of snails to infection.